To Explore The Mechanism Of Improving IGR By Ban Xia Xie Xin Tang Based On Insulin/GLP-1 Secretion Impairment And Clock Gene Level Disturbance Induced By 3DG In Rats

Objective:To investigate the mechanism of impaired glucose regulation(IGR)induced by 3-deoxyglucosone(3DG)in the circadian rhythm of insulin secretion mediated by biological clock genes,and the intervention effect of BanXia XieXin Decocion(BXXXT)on impaired glucose regulation induced by 3DG.Methods:1.In vivo experiment:The pre-diabetic rat model was established using 3DG.The successfully constructed rats were randomly divided into model group and experimental group,with 6 rats in each group,and 6 normal rats were selected as blank control group.The blank control group and model group were given 10 mL·kg-1·d-1 saline solution,while the experimental group(BXXXT group)were given 5 g·kg-1·d-1 for 14 days.After 14 days of intervention,oral glucose tolerance test(OGTT)was performed,in which 0 expression min glucose and insulin were used as fasting blood glucose and fasting insulin.Serum insulin secretion was determined by enzyme-linked immunosorbent assay(ELISA).The level of biological clock genes in rats was detected by reverse transcription polymerase chain reaction(RT-PCR).2.In vitro experiment:Islet β cell line INS-1 cells were used to model with 3DG(300 ng/mL).Banxia Xiexin Decoction containing serum to intervene.Insulin secretion of INS-1 cells was determined by ELISA.Results:1.a.In vivo experiment:(1)Compared with the control group,3DG group could significantly increase FBG,blood glucose and AUC of rats(P<0.01,P<0.05).(2)The OGTT curve of the control group reached the peak insulin secretion at the 30 min and returned to normal at the 120 min,which was consistent with the normal insulin secretion curve.The OGTT curve of 3DG group reached the peak at 60 min and recovered at 120 min.Compared with the control group,3DG group could significantly increase FINS(P<0.01),3DG group could significantly increase Serum insulin at 60 min(P<0.001),while the serum insulin level at 30 min was significantly decreased(P<0.001).(3)Compared with control group,3DG group could significantly increase insulin secretion of islets under low glucose condition(P<0.05);On the contrary,under the condition of high glucose,3DG group could significantly decrease the insulin secretion of the islet(P<0.05).(4)Compared with the control group,GLP-1 level in 3DG group showed a decreasing trend at 0min,30 min and 120 min,with a significant difference at 30 min(P<0.05),and an increasing trend at 90 min,but no significant statistical difference.(5)Compared with the control group,3DG group could significantly decrease the level of BMAL1 gene in colon tissue(P<0.01),3DG group could significantly increase the level of REV-ERBA gene(P<0.001),but there was no significant change in CLOCK gene.Compared with the control group,3DG group could not significant changes the levels of BMAL1 gene,CLOCK gene and REV-ERBA gene in the brain tissue.b.In vitro experiment:Compared with the control group,3DG group could significantly increase insulin secretion of INS-1 cells under the condition of low glucose(P<0.01),while 3DG group could significantly decrease insulin secretion of INS-1 cells under the condition of high glucose(P<0.01).2.a.In vivo experiment:(1)Compared with the control group,3DG group could significantly increase FBG,blood glucose and AUC of rats(P<0.01,P<0.05);Compared with 3DG group,BXXXT group could significantly decrease FBG,blood glucose and AUC of rats(P<0.001,P<0.01).(2)Compared with the control group,3DG group could significantly increase FINS(P<0.01),but the serum insulin level at 30 min was significantly decreased(P<0.001);Compared with 3DG group,BXXXT group could significantly decrease FINS(P<0.05),while serum insulin level at 30 min was significantly increased(P<0.001).(3)Compared with the control group,GLP-1 level in 3DG group showed a decreasing trend at 0 min,30 min and 120 min,with a significant difference at 30 min(P<0.05),and an increasing trend at 90 min,but no significant statistical difference.Compared with 3DG group,the GLP-1 level of BXXXT group showed an increasing trend at 0min,30 min,60 min and 120 min,with a significant difference at 30 min(P<0.05),and a decreasing trend at 90 min,but no significant difference.(5)Compared with the control group,3DG group could significantly decrease the level of BMAL1 gene in colon tissue of(P<0.01),and the level of REV-ERBA gene was significantly increased(P<0.001).However,there were no significant changes in the level of CLOCK gene and BMAL1 gene,CLOCK gene and REV-ERBA gene in rat brain tissue.Compared with 3DG group,BXXXT group could significantly increase BMAL1 gene level in colon tissue of(P<0.05),while REV-ERBA gene level was significantly decreased(P<0.05),there were no significant changes in the levels of CLOCK gene and BMAL1 gene,CLOCK gene and REV-ERBA gene in rat brain tissue.b.In vitro experiment:Compared with the control group,BXXXT group could significantly decrease insulin secretion of INS-1 cell under low glucose condition(P<0.001),while BXXXT group could significantly increase in insulin secretion of INS-1 cells under high glucose condition(P<0.001).(2)Compared with the control group,3DG group could significantly increase insulin secretion of INS-1 cells under the condition of low glucose(P<0.01),while 3DG group could significantly decrease insulin secretion of INS-1 cells under the condition of high glucose(P<0.01).Compared with the 3DG group,BXXXT group could significantly decrease insulin secretion of INS-1 cells under the condition of low glucose(P<0.05),while BXXXT group could significantly increase insulin secretion of INS-1 cells under the condition of high glucose(P<0.05).Conclusion:Based on the entero-pancreatic axis,3DG interferes with GLP-1 secretion by regulating the colonic clock gene,resulting in impaired insulin physiological secretion pattern,which is one of the mechanisms of impaired glucose regulation.Based on the entero-pancreatic axis,Banxia Xiexin Decoction can improve GLP-1 secretion dysfunction by regulating the colon clock gene and the impaired physiological secretion pattern of insulin,which may be one of the mechanisms by which it can improve the impaired glucose regulation induced by 3DG.Biological clock gene is the key regulatory factor of the imbalance of glucose metabolism homeostasis caused by the pathogenic factor 3DG in diet,and is also one of the key targets of Banxia Xiexin Decoction in the prevention and treatment of diabetes,which is expected to provide scientific basis for the mechanism research of diabetes and the prevention and treatment of diabetes with traditional Chinese medicine.